Acetyl-CoA is one of significantly important intermediates in metabolic pathways of microorganisms. Various metabolites are produced via acetyl-CoA. Well-known examples of such substances produced via acetyl-CoA include amino acids such as L-glutamic acid, L-glutamine, L-proline, L-arginine, L-leucine, and L-isoleucine; organic acids such as acetic acid, propionic acid, butyric acid, caproic acid, citric acid, 3-hydroxybutyric acid, 3-hydroxyisobutyric acid, 3-aminoisobutyric acid, 2-hydroxyisobutyric acid, methacrylic acid, and poly-3-hydroxybutyric acid; alcohols such as isopropyl alcohol, ethanol, and butanol; acetone; and polyglutamic acids.
In most microorganisms, acetyl-CoA is produced using a sugar such as glucose as a carbon source. The sugar is first converted into pyruvate via a metabolic pathway called the glycolytic pathway, such as the Embden-Meyerhof pathway, Entner-Doudoroff pathway, or the pentose phosphate pathway. Subsequently, the pyruvate is converted into acetyl-CoA by actions of decarboxylase, pyruvate formate-lyase, and the like. In this process, carbon dioxide and formate are generated as byproducts, and some of the carbons derived from the sugar will lost. Therefore, several studies have been carried out with the aim of achieving re-fixation of carbon dioxide in order to increase the yield of acetyl-CoA.
In microorganisms, there are several known pathways for fixing carbon dioxide as a carbon source (Appl. Environ. Microbiol. 77(6), 1925-1936, 2011). Specific examples of the pathways include the Calvin-Benson cycle, the reductive TCA cycle, the Wood-Ljungdahl pathway, the 3-hydroxypropionate cycle, and the 4-hydroxybutyrate cycle. The Calvin-Benson cycle is a CO2 fixation pathway existing in plants and photosynthetic bacteria, and containing about 12 kinds of enzymes. In the Calvin-Benson cycle, CO2 is fixed by ribulose-1,5-bisphosphate carboxylase (RubisCO) and, ultimately, glyceraldehyde 3-phosphate is produced. The reductive TCA cycle is found in microaerophilic bacteria and anaerobic bacteria including green sulfur bacteria, and contains 11 kinds of enzymes. This cycle is characterized by CO2 fixation enzymes (i.e., acetyl-CoA carboxylase, 2-oxoglutarate synthase) that requires ferredoxin as a coenzyme. In the reductive TCA cycle, pyruvate is produced from CO2 by the reverse reaction of the usual TCA cycle. The Wood-Ljungdahl pathway is found in anaerobic microorganisms such as acetic acid-producing bacteria, and contains 9 kinds of enzymes. In the Wood-Ljungdahl pathway, CO2 and formate on a coenzyme are reduced by formate dehydrogenase, CO dehydrogenase, etc., and, ultimately converted into acetyl-CoA. The 3-hydroxypropionate cycle is found in Chloroflexus bacteria and the like, and contains 13 kinds of enzymes. In the 3-hydroxypropionate cycle, CO2 is fixed by the action of acetyl-CoA (propionyl-CoA) carboxylase and acetyl-CoA is produced via malonyl-CoA and the like. The 4-hydroxybutyrate cycle exists in archaeabacteria and the like. In the 4-hydroxybutyrate cycle, CO2 is fixed by the actions of pyruvate synthase, acetyl-CoA (propionyl-CoA) carboxylase, and phosphoenolpyruvate carboxylase, whereby acetyl-CoA is produced via 4-hydroxybutyryl CoA and the like.
In order to produce a useful substance, several approaches have been reported as ideas to introduce a carbon dioxide fixation pathway to a useful-compound-producing microorganism. For example, International Publication (WO) 2009/094485 and WO 2010/071697 disclose approaches to producing acetyl-CoA from carbon dioxide, by using a microorganism to which a pathway similar to the Wood-Ljungdahl pathway of acetic acid bacteria was introduced. As an example of CO2 fixation for producing a useful compound, WO 2009/046929 discloses an approach to producing lactic acid from carbon dioxide by using a microorganism to which hydrogenase and tetrahydrofolate lyase were introduced. WO 2011/099006 proposes a cycle in which CO2 is fixed via a carbon dioxide fixation reaction onto acetyl-CoA or a malonyl-CoA reduction reaction. German Patent No. 102007059248 proposes production of acetyl-CoA by a pathway similar to the 4-hydroxybutyrate cycle.